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Mining enzymes from extreme
environments
熊玉翠
10708054
What is Metagenomics?
Metagenomics is a strategy, characterizing DNA extracted directly from
environmental samples.
Current advances in metagenomics have revolutionized the research in
fields of microbial ecology and biotechnology, enabling the highthroughput discovery of new enzymes for industrial bioconversions.
南极冰下湖存在着复杂的微生物群
美國黃石公園的熱泉內棲息著地球上最原
始的微生物
南极深处发现的活着的微生物
Two obstacles :
First, most extremophiles cannot be cultured using traditional culturing
technologies
Second, the very low biomass densities often do not yield enough DNA
and reduces the effectiveness of cloning
Introduction：
The extremophiles thrive in the harshest environments.
Methanococcus jannaschii
Currently, about 4% of all sequencing projects deal with extremophiles
that have industrial applications.
however, if the final goal is the identification of extremozymes
we must also improve annotations through empirical work on
specific candidates
not all proteins from extreme environments are
extremozymes
First, in some environments(extreme temperature orpressure)
they are expected to be extremozymes
both intracellular and extracellular biochemical machineries are therefore
exquisitely adapted to such conditions.
Second, in other environments(extremes of pH or salinity/osmolarity,
the presence of organic solvents or high levels of radiation)
they are not expected to be extremozymes
maintain intracellular conditions more typical of non-extremophiles
Estimates of microbial diversity in extreme and
non-extreme microbial communities
The microbial diversity on our planet is mostly comprised by the
microorganisms, in terms of both absolute numbers of cells and of the
biomass, and the numbers of species.
Different culture-independent technologies
Recent data suggests that the community composition and diversity can
shift greatly in human-perturbed areas, as well as within small vertical
and horizontal distances, possibly in response to changes in the
extreme physical and chemical conditions.
Protein engineering versus metagenomics
In the past two years, 45 extremozymes have been identified from
pure cultures isolated from different extreme environments.
Although, many of these enzymes have been proven to be useful for
chemical synthesis, one still might exhibit some unpredictable
features with unnatural substrates and often exhibit low operational
stability.
To overcome these problems…
using protein engineering and chemical modification screening it
is currently possible to create ‘artificial extremozymes’ with
altered protein topology, thermal stability and tolerance to
organic solvents.
However, these strategies have a few hindrances, because the
accumulation of subtle changes at genetic level do not always
lead to the best enzymatic fitness.
Metagenomics constitutes a challenging researchdomain with great
potential for both fundamental and industrial applications that range
from the understanding of microbial adaptation and evolution to the
discovery of new enzymes for their direct use.
challenges and bottlenecks
One of the main challenges for mining enzymes from extreme
environments is not only to develop a robust screening or selection
system to select for specific extremozymes, but also to overcome
the problem of the low biomass yields and very low cell numbers that
hinder high yields of DNA for cloning.
To solve this problem…
multiple displacement amplification (MDA)
amplification(WGA)
whole genome
high-throughput sequencing has revolutionized the study of microbial
community metagenomes
certain limitations and difficulties
Prospect
Future advances should focus on designing cell-free systems with a
double aim:
to increase substrate bioavailability and to reduce inhibition and cross
reactivity of cell components.
Thanks for your attention!
DGGE电泳设备是用于进行DGGE电泳的仪器，
DGGE技术是DNA指纹技术。
定量PCR
FISH荧光原位杂交
1) The random hexamers (represented by a blue line) bind to the denatured DNA
(represented by a green line); (2) The DNA polymerase (represented by a blue
circle) extends the primers until it reaches newly synthesized double-stranded
DNA (represented by an orange line); (3) The enzyme proceeds to displace the
strand and continues the polymerization, while primers bind to the newly
synthesized DNA; (4) Polymerization starts on the new strands, forming a
hyperbranched structure.
(
Applied Biosystems Prism 3700型基因分析仪
为96道毛细管基因分析仪，是高通量基因分析
的技术平台。
超高通量基因组测序系统GS FLX 系统